Abuse and addiction to prescription opioids (short-acting MOP-r agonists such as oxycodone) have reached epidemic proportions in the US. Therapeutic options to block the progression of trajectory from abuse to addiction are limited. The concept of vaccination against drugs of abuse has therefore emerged for such a preventive purpose. However small molecules such as drugs of abuse are not highly immunogenic, therefore standard approaches include conjugation of the antigen epitope (e.g., oxycodone) to a protein immunogen. Effective vaccination is postulated to result in sequestration (and thus inactivation) of the drug of abuse in the bloodstream, through the action of a sufficient population of high-affinity antibodies. However, standard approaches have a number of technical limitations that can limit their reliability and effectiveness. The goal of this CEBRA proposal is therefore to develop a mechanistically novel vaccination platform based on the orderly and high-density presentation of antigen epitopes in the coat of the extracellular protozoan parasite Trypanosoma brucei (T. brucei). This biologically-evolved antigen-presentation mechanism results in strong B-cell responses and B-cell memory, crucial to effective vaccination. Aim 1 of this multi- disciplinary proposal will therefore develop the first . brucei platform for small molecule vaccination, utilizing orderly presentation of oxycodone antigen moieties to decorate sortase tags transgenically expressed on the coat of T. brucei. Vaccination with inactivated T. brucei (having no potential for pathogenicity) will then be modeled in rats, and the production and time course of selective anti-oxycodone antibodies will be optimized. Aim 2 will then focus on the proof-of-technology validation of this approach. This will be done through optimization of the effectiveness of anti-oxycodone vaccination in rats, monitored by pharmacokinetic (e.g., decrease in blood oxycodone levels) and pharmacodynamic (blockade of oxycodone-induced analgesia and of i.v. oxycodone self-administration) parameters. The development of this cutting-edge vaccination strategy for abused small molecules (with oxycodone as a prototype) therefore has considerable future potential for clinical translation, and for drug abuse research.